Solid-phase peptide synthesis (SPPS) has become a mainstay method for the synthesis of small to medium length peptides and proteins. Peptides of substantially longer length can be prepared using SPPS methods owing to the advent of native chemical ligation methods. See Merrifield, R. B. J. Am. Chem. Soc. 1963, 85, 2149 and Dawson, P. E.; Muir, T. W.; Clarklewis, I.; Kent, S. B. H. Science 1994, 266, 776-779. However, removal of deletion sequences, that often accumulate during synthesis, from the desired full-length product remains a tedious, undesirable aspect of SPPS. A similar problem has also been noted in the solid-phase synthesis of oligosaccharides. See P. H. Seeberger et al. Angew. Chem. Int. Ed. 2001, 40, 4433 and U.S. Patent Application 2003/0232978. In particular, removal of sequences, e.g., peptide or oligosaccharide sequences, differing by only one unit (so-called “n−1 products”), which stem from incomplete conversion at any stage of the synthesis, can be very difficult. In response to the need for better purification procedures, a number of methods have been reported for the isolation and purification of biopolymers, including biopolymers produced by chemical synthesis or recombinant DNA techniques. For example, centrifugation, column chromatography, and electrophoresis can be used to purify biopolymers. Unfortunately, these methods require one or more additional and often burdensome purification steps after initial purification of the biopolymer. These purification procedures are also unsatisfactory because a significant amount of the crude biopolymer is often lost during the procedure resulting in reduced yields.
To facilitate purification of peptides and proteins prepared by SPPS, various reagents have been developed with the intention of tagging deletion sequences or the full-length product. The compound bearing the tag has different chemical properties and can be separated from other compounds using various purification methods, e.g., chromatographic purification. Several chemical groups that function as purification handles have been described that take advantage of selective conditions under which the retention time can be varied on ion-exchange, hydrophobic interaction, affinity and other types of chromatographic columns. See Funakoshi, S.; Fukuda, H.; Fujii, N., J. Chromatogr. 1993, 638, 21-27; Shogren-Knaak, M. A.; McDonnell, K. A.; Imperiali, B., Tetrahedron Lett. 2000, 41, 827-829; Shogren-Knaak, M. A.; Imperiali, B. Tetrahedron Lett. 1998, 39, 8241-8244; Villain, M.; Vizzavona, J.; Rose, K. Chem. Biol. 2001, 8, 673-679; Vizzavona, J.; Villain, M.; Rose, K. Tetrahedron Lett. 2002, 43, 8693-8696; Canne, L. E.; Winston, R. L.; Kent, S. B. H. Tetrahedron Lett. 1997, 38, 3361-3364; and P. C. de Visser et al. Tetrahedron Lett. 2003, 9013. Nevertheless, purification of peptides remains a difficult task, particularly purification of peptide sequences differing by only one residue (n−1 products). For additional discussion relating to solid-phase peptide synthesis see Cotton, G. J.; Muir, T. W. Chem. Biol. 1999, 6, R247-R256; Canne, L. E.; Botti, P.; Simon, R. J.; Chen, Y. J.; Dennis, E. A.; Kent, S. B. H. J. Am. Chem. Soc. 1999, 121, 8720-8727; Dawson, P. E.; Kent, S. B. H. Annu. Rev. Biochem. 2000, 69, 923-960; and Kochendoerfer, G. G.; Kent, S. B. H. Curr. Opin. Chem. Biol. 1999, 3, 665-671.
Fluorous-phase purification is an attractive technique for the purification of peptides, oligosaccharides, oligonucleotides, and small organic compounds. In fluorous purification procedures, prior to purification a group containing a relatively large number of fluorine atoms is attached to the compound to be purified. The step of attaching the fluorous group to the compound imparts unique chemical properties to the compound, thereby making it easier to separate the fluoro-tagged compound from other compounds. In general, fluorous-tagged molecules partition preferentially into a fluorous phase while non-tagged molecules partition into an organic phase. The fluorous tag can be installed while the compound is attached to the solid phase or the fluorous tag can be installed using solution-phase chemistry. Recently, highly-fluorinated compounds have been employed in protein design, reaction acceleration catalysis, combinatorial chemistry, and organic separation methodology. Among the canonical amino acid side chain functionalities, none are expected to extensively interact with fluorous materials. Thus, an appendage that is highly fluorinated should be unique in its physical properties, compared to most peptide products. See Filippov, D. V.; van Zoelen, D. J.; Oldfield, S. P.; van der Marel, G. A.; Overkleeft, H. S.; Drijfhout, J. W.; van Boom, J. H. Tetrahedron Lett. 2002, 43, 7809-7812. Although fluorous synthetic and/or separation techniques are promising, such techniques are currently limited by a lack of available and suitable fluorinated compounds that can be used to install fluorinated molecular tags.
Therefore, the need exists for fluorinated compounds that can be used to install fluorinated molecular tags and methods of purification using the fluorinated molecular tags. The present invention fulfills this need and has other related advantages.